In vitro research involving horses refers to the study of equine cells, tissues, or biological molecules outside their normal biological context, typically in controlled laboratory environments. This research approach allows scientists to investigate cellular processes, molecular interactions, and the effects of various treatments without the ethical and logistical complexities of in vivo studies. In vitro studies contribute to understanding equine physiology, pathology, and pharmacology by providing insights into cellular responses to pathogens, drugs, and other stimuli. This page compiles peer-reviewed research studies and scholarly articles that explore various in vitro methodologies and their applications in equine science, including cell culture techniques, molecular assays, and drug efficacy testing.
Moore AI, Squires EL, Graham JK.Seminal plasma is generally removed from equine spermatozoa prior to cryopreservation. Two experiments were designed to determine if adding seminal plasma back to spermatozoa, prior to cryopreservation, would benefit the spermatozoa. Experiment 1 determined if different concentrations of seminal plasma affected post-thaw sperm motility, viability and acrosomal integrity of frozen/thawed stallion spermatozoa. Semen was washed through 15% Percoll to remove seminal plasma and spermatozoa resuspended to 350 x 10(6)sperm/mL in a clear Hepes buffered diluent containing either 0, 5, 10, 20, 40 or 80%...
Rambags BP, Stout TA.An 18-year-old Friesian mare with a large intrauterine cyst was examined by transrectal ultrasonography. There were several small to moderately sized intraluminal endometrial cysts, one of which connected via the myometrium to a large subserosal cyst, thereby effectively forming a 'transmural' cyst complex. During a videohysteroscopy, the intraluminal part of this transmural cyst was removed by electrocoagulation via a polypectomy snare. It was then possible to drain the large subserosal part of the cyst into the uterine lumen by transrectal massage, thereby confirming the presence of the tran...
Moussa M, Bersinger I, Doligez P, Guignot F, Duchamp G, Vidament M, Mermillod P, Bruyas JF.Vitrification using open pulled straw (OPS) has provided encouraging results with embryos from other species. The aim of this study was to compare the survival of 6.5- and 6.75-day-old equine embryos after OPS vitrification and slow-cooling. Eighteen embryos were frozen using a slow-cooling method. Embryos were placed in modified PBS with increasing glycerol concentration (2.5%, 5%, 7.5% and 10% (v/v) 5 min each). Embryos were loaded into 0.25 ml straws then placed in a programmable freezer and subsequently plunged into liquid nitrogen. After thawing, cryoprotectant was removed by five steps w...
The measurement of cell proliferation and cell viability using 5'bromo-2'deoxy-uridine (BrdU) labelling has been described in several cell types and species. The aim of this study was to adapt this technique to equine embryos and to compare the index of DNA replication (S-phase) between equine and caprine embryos. Seventeen equine embryos were recovered at day 6.5 post-ovulation and 20 caprine embryos were recovered at day 7 after the onset of estrus. Equine embryos were incubated during 1h at 39 degrees C in PBS containing 1mM of BrdU. Embryos were then treated in 0.05% trypsin during 15 min ...
Taylor DD, Hood DM, Potter GD, Hogan HA, Honnas CM.To evaluate patterns of digital cushion (DC) displacement that occur in response to vertical loading of the distal portion of the forelimb in horses. Sample Population-Forelimbs from 10 horses with normal feet. Methods: Patterns of DC displacement induced by in vitro vertical limb loading were determined. Load-induced displacement of the DC was defined as the magnitude and direction of displacement of 6 radio-dense, percutaneously implanted markers in specific regions of the DC. The effects of solar support and nonsupport on displacement of the DC were compared. Results: Regional displacement ...
Dechant JE, Baxter GM, Frisbie DD, Trotter GW, McIlwraith CW.Clinical trials in human and veterinary literature have documented the benefits of oral nutraceutical joint supplements containing glucosamine (GU) and chondroitin sulphate (CS) to treat mild to moderate osteoarthritis, but the effects of these components have not yet been conclusively determined. Objective: To assess varying dosages of GU and CS on normal and interleukin-1alpha (IL-1) conditioned equine cartilage explants and rationalise the use of these products. Objective: Treatment would not be detrimental to cartilage metabolism and higher dosages and the combination of GU and CS would be...
Akens MK, Hurtig MB.Bovine articular cartilage is often used to study chondrocytes in vitro. It is difficult to correlate in vitro studies using bovine chondrocytes with in vivo studies using other species such as rabbits and sheep. The aim of this investigation was to study the effect of species, anatomical location and exogenous growth factors on chondrocyte proliferation in vitro. Methods: Equine (EQ), bovine (BO) and ovine (OV) articular chondrocytes from metacarpophalangeal (fetlock (F)), shoulder (S) and knee (K) joints were cultured in tissue culture flasks. Growth factors (rh-FGFb: 10 ng/ml; rh-TGFbeta: 5...
Sankaranarayanan R, Biswal BK, Vijayan M.A new relaxed state has been characterized in the crystals of horse methemoglobin grown at neutral pH at low ionic concentration and their low humidity variants. The crystals provide an example for improvement in X-ray diffraction quality with reduced solvent content. Only the classical R state has been so far observed in liganded horse hemoglobin. The state characterized in the present study lies in between the R state and the R2 state characterized earlier in liganded human hemoglobin. The results presented here, along with those of earlier studies, suggest that relaxed and tense hemoglobin ...
Sepulveda MF, Greenaway EC, Avella M, Goode NT, Cunningham FM.To determine if protein kinase C (PKC) regulates equine eosinophil function. Methods: Blood eosinophils were obtained from healthy ponies. Methods: IL-5- and histamine-induced adherence to serum-coated plastic was measured as the eosinophil peroxidase content of adherent cells and serum treated zymosan (STZ)-and IL-5-induced superoxide production by the reduction of cytochrome C. Eosinophil PKC activity was quantitated as the rate of transfer of (32)P from ATP to substrate. The effects of Ro31-8220 (isotype non-selective PKC inhibitor), Go6976 (conventional PKC inhibitor), and rottlerin (PKCde...
Spencer JA, Deinnocentes P, Moyana EM, Guarino AJ, Ellison SE, Bird RC, Blagburn BL.Equine protozoal myeloencephalitis (EPM) is a neurologic syndrome seen in horses from the Americas and is mainly caused by Sarcocystis neurona. Recently, a 29-kDa surface antigen from S. neurona merozoites was identified as being highly immunodominant on a Western blot. This antigen has been sequenced and cloned, and the expressed protein has been named SnSAG1. In a previous study, cell-mediated immune responses to SnSAG1 were shown to be statistically significantly reduced in horses with EPM in comparison to EPM-negative control horses. It therefore appears as though the parasite is able to i...
Andritzky J, Rossol M, Lischer C, Auer JA.To compare the precision obtained with computer-assisted screw insertion for treatment of mid-sagittal articular fractures of the distal phalanx (P3) with results achieved with a conventional technique. Methods: In vitro experimental study. Methods: Thirty-two cadaveric equine limbs. Methods: Four groups of 8 limbs were studied. Either 1 or 2 screws were inserted perpendicular to an imaginary axial fracture of P3 using computer-assisted surgery (CAS) or conventional technique. Screw insertion time, predetermined screw length, inserted screw length, fit of the screw, and errors in placement wer...
Watson ED, Bae SE, Al-Zi'abi MO, Hogg CO, Armstrong DG.The insulin-like growth factors, IGF-I and -II, have been shown to play a key role in luteal function in some species. The IGF binding proteins, IGFBP-2 and -3, have been shown to inhibit binding of IGF-I and -II to bovine luteal cells and decrease progesterone production. We have recently shown that equine follicles have the genetic capacity to produce IGFBP-2, and that levels decrease in healthy preovulatory follicles. In the present study expression of mRNAs encoding IGFBP-2, as well as the rate-limiting steroidogenic enzyme, P450scc, were studied in equine corpora lutea to investigate whet...
Lafont R, Dinan L.Ecdysteroids are widely used as inducers for gene-switch systems based on insect ecdysteroid receptors and genes of interest placed under the control of ecdysteroid-response elements. We review here these systems, which are currently mainly used in vitro with cultured cells in order to analyse the role of a wide array of genes, but which are expected to represent the basis for future gene therapy strategies. Such developments raise several questions, which are addressed in detail. First, the metabolic fate of ecdysteroids in mammals, including humans, is only poorly known, and the rapid catabo...
Traversa D, Otranto D, Iorio R, Giangaspero A.Equine thelaziosis caused by the eyeworm Thelazia lacrymalis is a parasitic disease transmitted by muscid flies. Although equine thelaziosis is known to have worldwide distribution, information on the epidemiology and presence of the intermediate hosts of T. lacrymalis is lacking. In the present work, a PCR-RFLP based assay on the first and/or second internal transcribed spacer (ITS1 and ITS2) of ribosomal DNA was developed for the detection of T. lacrymalis DNA in its putative vector(s). The sensitivity of the technique was also assessed. The restriction patterns obtained readily differentiat...
Maury W, Thompson RJ, Jones Q, Bradley S, Denke T, Baccam P, Smazik M, Oaks JL.Equine infectious anemia virus (EIAV) is a lentivirus with in vivo cell tropism primarily for tissue macrophages; however, in vitro the virus can be adapted to fibroblasts and other cell types. Tropism adaptation is associated with both envelope and long terminal repeat (LTR) changes, and findings strongly suggest that these regions of the genome influence cell tropism and virulence. Furthermore, high levels of genetic variation have been well documented in both of these genomic regions. However, specific EIAV nucleotide or amino acid changes that are responsible for cell tropism changes have ...
McConnico RS, Stokes AM, Eades SC, Moore RM.To examine the secretory response (in the presence and absence of prostaglandin inhibition) in vitro and structural alterations of colonic mucosa in horses after intragastric administration of black walnut extract (BWE). Methods: 14 adult horses. Methods: Seven horses were administered BWE intragastrically and monitored for 11 hours. Tissue samples were obtained from the right ventral, left ventral, and right dorsal colons (RVC, LVC, and RDC, respectively) of the 7 BWE-treated and 7 control horses. Tissue samples were examined via light microscopy, and the extent of hemorrhage, edema, and gran...
Kirk ES, Squires EL, Graham JK.Assessing the fertilizing potential of a semen sample is important for effective stallion management and for rapid progress in evaluating new cryopreservation technologies. Unfortunately, sperm motility does not estimate fertility well. These experiments established assays to measure cell viability, acrosomal integrity and mitochondrial function for cryopreserved stallion spermatozoa, using flow cytometry, and determined the variability associated with these assays. Correlations between results for these laboratory assays and stallion fertility were also determined. The inter-assay variability...
Aziz DM, Ahlswede L, Enbergs H.The assay of MTT reduction depends on the ability of metabolically active cells to reduce the tetrazolium salt (3[4,5-dimethylthiazol-2-y1]-2,5-diphenyltetrazolium bromide) to formazan. This study was conducted to examine and validate of a simple and less costly MTT test in determining equine sperm viability and compare the efficiency of this test with a flow cytometer. Fresh ejaculates from 11 stallions (warm blood) were included in this study. Semen was diluted to 100 million cells/ml in a Hepes 0.1% BSA. The rates of MTT reduction were measured in microtiter plates after incubation for 1 an...
Mobasheri A, Gent TC, Womack MD, Carter SD, Clegg PD, Barrett-Jolley R.In this comparative study, we have established in vitro models of equine and elephant articular chondrocytes, examined their basic morphology, and characterized the biophysical properties of their primary voltage-gated potassium channel (Kv) currents. Using whole cell patch-clamp electrophysiological recording from first-expansion and first-passage cells, we measured a maximum Kv conductance of 0.15 +/- 0.04 pS/pF (n = 10) in equine chondrocytes, whereas that in elephant chondrocytes was significantly larger (0.8 +/- 0.4 pS/pF, n = 4, P </= 0.05). Steady-state activation parameters of eleph...
Wagner B, Robeson J, McCracken M, Wattrang E, Antczak DF.Recombinant cytokines are valuable tools for functional studies and candidates for vaccine additives or therapeutic use in various diseases. They can also be used to generate specific antibodies to analyze the roles of different cytokines during immune responses. We generated a mammalian expression system for recombinant cytokines using the equine IgG1 heavy chain constant region as a tag for detection and purification of the expressed cytokine, demonstrated here using equine interferon-gamma (IFN-gamma), interleukin-2 (IL-2), interleukin-4 (IL4) and transforming growth factor-beta1 (TGF-beta1...
Steenkamp G, Olivier-Carstens A, van Heerden WF, Crossley DA, Boy SC.Surgical endodontic therapy is a conservative dental technique used in horses with some degree of clinical success. Failure of this procedure can partially be explained by inadequate sealing of the root apices with resultant microleakage in the periapical area. Objective: To assess and compare in vitro sealing ability of 3 different dental restorative materials used as apical sealants during equine surgical endodontics. Methods: Thirty extracted equine cheek teeth were divided randomly into 3 groups and subjected to apicoectomy and apical sealing using 3 materials: reinforced zinc oxide-eugeno...
Lamikanra A, Myers KA, Ferris N, Mitrophanous KA, Carroll MW.Lentiviral-based vectors hold great promise as gene delivery vehicles for the treatment of a wide variety of diseases. We have previously reported the development of a nonprimate lentiviral vector system based on the equine infectious anaemia virus (EIAV), which is able to efficiently transduce dividing and nondividing cells both in vitro and in vivo. Here, we report on the application of EIAV vectors for the systemic delivery of an antibody fusion protein designed for the treatment of cancer. The therapeutic potential of a single chain antibody against the tumour-associated antigen, 5T4, fuse...
Love CC, Brinsko SP, Rigby SL, Thompson JA, Blanchard TL, Varner DD.The relationship between seminal plasma level (0, 10, or 20%) and extender type [Kenney type (EZ-Mixin-CST) or Kenney-modified Tyrodes-KMT] to the susceptibility of sperm DNA to denaturation and sperm motility measures were investigated in cooled (5 degrees C) stallion sperm. Three ejaculates from each of three fertile stallions were collected in an artificial vagina and processed as follows: diluted one part uncentrifuged semen with four parts of extender to a final concentration of 20% seminal plasma in either CST or KMT (20% CST; 20% KMT); diluted to a final concentration of 25 million sper...
de la Fuente J, Ayoubi P, Blouin EF, Almazán C, Naranjo V, Kocan KM.Anaplasma phagocytophilum (Rickettsiales: Anaplasmataceae) causes human, equine and canine granulocytic anaplasmosis and tick-borne fever of ruminants. The rickettsia parasitizes granulocytes and bone marrow progenitor cells, and can be propagated in human promyelocytic and tick cell lines. In this study, microarrays of synthetic polynucleotides of 21,329 human genes were used to identify genes that are differentially expressed in HL-60 human promyelocytic cells in response to infection with A. phagocytophilum. Semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) of selec...
Nagata LP, Hu WG, Masri SA, Rayner GA, Schmaltz FL, Das D, Wu J, Long MC, Chan C, Proll D, Jager S, Jebailey L, Suresh MR, Wong JP.The efficacy of a DNA vaccine against western equine encephalitis (WEE) infection in mice was evaluated. The 26S structural region was expressed, in vitro from an internal T7 promoter using a rabbit reticulysate transcription/translation system; and from a CMV promoter after transfection into Vero cell monolayers. The proteins synthesized were reactive with anti-WEE virus (WEEV) antibodies, both in western blot analysis and histochemical staining, respectively. When the DNA vaccine plasmid, pVHX-6, was administered intraepidermally to mice, followed by challenge in a lethal mouse model, the le...
Grignard E, Morin J, Vernet P, Drevet JR.We report here on the cloning of cDNAs coding bovine and equine orthologs of mouse epididymis-restricted and sperm-bound glutathione peroxidase 5 (GPX5), a selenium-independent member of the multigenic GPX family in mammals. The complete sequence of bovine GPX5 as well as a partial sequence of the equine GPX5 were characterized, conceptually translated and aligned with other known mammalian GPX5 proteins. Using Northern blotting assays, we show that the level of expression of GPX5 is high in bovine but low in equine and that in both species the regionalization of GPX5 expression in epididymis ...
Zerpa H, Vega F, Vasquez J, Ascanio E, Campos G, Sogbe E, Romero E, Ascanio M, García H.Endotoxaemia is a syndrome linked to the development of equine laminitis; however, the relationship between them is uncertain. The aim of this experiment was to evaluate the effect of an experimental acute sublethal endotoxaemia model on in vitro equine palmar digital vascular reactivity. Rings of arteries and veins of each forelimb were obtained from 11 clinically healthy horses submitted to two surgical procedures, 3 weeks apart. Before the second surgery, 0.25 microg/kg of lipopolysaccharide from Escherichia coli O55:B5 in saline, was administered i.v. in 30 min. After 3 h, the vessels were...
van der Harst MR, Bull S, Laffont CM, Klein WR.The aminoglycoside gentamicin is often used in equine practice. Despite its clinical use, concerns remain regarding the potential toxic side-effects, such as nephrotoxicity, in equine patients, particularly after repeated dosing. The aim of the study was to investigate first in vitro the mechanisms contributing to the renal toxicity of gentamicin and to identify sensitive biomarkers indicating proximal tubule damage. To this end, the kidney-derived cell lines LLC-PKI and MDCK were treated with gentamicin at different concentrations. Toxicity was assessed by measuring the release of gamma-gluta...
Suh TK, Schenk JL, Seidel GE.Sexing sperm by high-speed flow cytometry subjects them to high pressure. The routine operating pressure of the MoFlo SX flow cytometer for sperm sorting for commercial production has been 50 pounds/square inch (psi), with a standard 70 microm standard nozzle tip. It was hypothesized that lowering the sorting pressure could reduce sperm damage. Therefore, a series of experiments using semen from six bulls, sorted with three MoFlo SX sorters, was conducted to determine optimal pressure. An additional experiment was done with stallion spermatozoa. In Experiment 1, sorting at 30 psi compared to 5...
Petrov R, MacDonald MH, Tesch AM, Benton HP.To investigate the effect of adenosine kinase inhibition on interleukin (IL)-1beta- and lipopolysaccharide (LPS)-induced cartilage damage. Methods: Articular cartilage was obtained from the metacarpophalangeal joints of 10 young adult horses. Following a stabilization period, weighed cartilage explants were exposed to IL-1beta (10 ng/ml) or LPS (50 microg/ml) to induce cartilage degradation. To test the potential protective effects of adenosine, these explants were simultaneously exposed to adenosine (100 microM), the adenosine kinase inhibitor 5'iodotubercidin (ITU, 1 microM) or to both adeno...
Kumar R, Prabhu NP, Bhuyan AK.Laser flash photolysis and stopped-flow methods have been used to study the dynamic events in the micro- to millisecond time bin in the refolding of horse ferrocytochrome c in the full range of guanidine hydrochloride concentration at pH 12.8 (+/-0.1), 22 degrees C. Under the absolute refolding condition, the earliest relaxation time of the unfolded protein chain is less than 1 micros. The chain then undergoes diffusive dynamics-mediated contraction and expansion, in which intrapolypeptide ligands make transient contacts with the heme iron, giving rise to two distinct kinetic phases of approxi...
Zink MC, Johnson JA, Prescot JF, Pascoe PJ.The in-vitro interaction of Corynebacterium equi and foal alveolar macrophages was examined qualitatively and quantitatively using cells collected by sequential bronchoalveolar lavage at 2-week intervals from birth until 14 weeks of age. Total and differential counts were performed on the recovered cells. Macrophages were identified using the non-specific esterase strain. Cultures of the alveolar macrophages were challenged with C. equi suspensions and the process and extent of ingestion was examined by light and electron microscopy. Few macrophages were recovered from the lungs of foals less ...
Cochrane CA, Freeman KL, Knottenbelt DC.Wound healing in equidae is delayed and more complicated than in other species. These complications arise from a condition known as exuberant granulation tissue formation. The lower limb of the horse is frequently slower to heal than other parts of the body and has a particular tendency to produce excess (exuberant) granulation tissue. Sarcoids are tumor-like lesions of the skin which often appear at the site of wounds. This study compared the growth characteristics of the sarcoid and granulation tissue-derived cells with normal dermal fibroblasts grown from primary cell cultures. All three ce...
Sanny CG, Weiner H.The inhibition of mitochondrial (pI 5) horse liver aldehyde dehydrogenase by disulfiram (tetraethylthiuram disulphide) was investigated to determine if the drug was an active-site-directed inhibitor. Stoichiometry of inhibition was determined by using an analogue, [35S]tetramethylthiuram disulphide. A 50% loss of the dehydrogenase activity was observed when only one site per tetrameric enzyme was modified, and complete inactivation was not obtained even after seven sites per tetramer were modified. Modification of only two sites accounted for a loss of 75% of the initial catalytic activity. Th...
Thomas PG, Ball BA, Miller PG, Brinsko SP, Southwood L.Attachment of spermatozoa to oviductal epithelial cells (OEC) may be a prefertilization event in some species. We tested the hypothesis that spermatozoa that attach to equine OEC monolayers are a selected subpopulation of the initial inseminate, containing a higher proportion of morphologically normal, motile cells than the inseminate. Washed stallion spermatozoa were cocultured with monolayers of OEC or monolayers of Vero cells, and controls were incubated in wells coated with basement membrane extract (Matrigel [Mgel]) or in plastic (uncoated) wells. Unattached spermatozoa were removed by ri...
Gimenez F, Hines SA, Evanoff R, Ojo KK, Van Voorhis WC, Maly DJ, Vidadala RSR, Mealey RH.Theileria equi, an etiologic agent of equine piroplasmosis, is a tick-transmitted hemoprotozoan of the phylum Apicomplexa. Recent outbreaks of piroplasmosis in the United States have renewed interest in safe and effective treatment options. Although imidocarb dipropionate (IMD) is the drug of choice for clearance of T. equi, adverse reactions and recently documented resistance support the need for alternative therapeutic strategies. The recently described bumped kinase inhibitors (BKIs) are a new class of compounds that could potentially be used as safe and effective alternatives to IMD. In an...
Matoba H, Allen JR, Bayly WM, Oakley CR, Gollnick PD.Comparisons were made of the histochemical characteristics of skeletal muscle from 10 animal species. The basic comparison was made from the staining patterns for the myofibrillar actomyosin ATPase produced by preincubation of fresh frozen cross-sections of muscle at alkaline pH (10.30) or acid pH (4.60) with those produced by preincubation in media containing Cu2+ at alkaline pH (10.30), near neutral pH (7.40), or acid pH (4.60). Muscle sections were also stained for reduced nicotinamide adenine dinucleotide tetrazolium reductase and alpha-glycerophosphate dehydrogenase to provide an indicati...
Quentin-Froignant C, Kappler-Gratias S, Top S, Bertagnoli S, Gallardo F.Equine herpesvirus 1 (EHV-1) is a causative agent of respiratory disorders, abortion and myeloencephalopathy in horses and has an important impact on equine health and economy. Several bacterial artificial chromosomes have already been developed and enabled identification and functional characterization of EHV-1 genes. Unfortunately, little is known about its replication. Here, the ANCHOR system was inserted by targeted homologous recombination into the equine herpesvirus genome. This insertion led to the conversion of EHV-1 DNA to auto-fluorescent spots easily detectable by fluorescence micro...
Pintado CO, Friend M, Llanes D.This paper describes two anti-glycoprotein IIb/IIIa or CD41/61 murine monoclonal antibodies (Co.35E4 and Co.2oA1). The cellular distribution and apparent molecular weight of the antigen detected by these antibodies is consistent with their reaction with ruminant and equine glycoprotein IIb/IIIa. Biochemical analysis of the equine molecule using sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) revealed bands of 24, 100 and 110 kDa under reducing conditions and 115 and 80 kDa under nonreducing conditions. Biochemical analysis of ruminant antigen revealed that the 24 kDa band...
Peterman BF, Morton RA.The apparent equilibrium constant and rate of oxidation was investigated for the reaction of cytochrome c with iron hexacyanide. It was found that if horse heart ferricytochrome c was exposed to ferricyanide (to oxidize traces of reduced protein) the cytochrome subsequently, even after extensive dialysis, had an apparent equilibrium constant different from that of electrodialyzed protein. The effect of ferricyanide ion apparently cannot be removed by ordinary dialysis. The ionic strength dependence of the apparent equilibrium constant and bimolecular oxidation rate constant was measured in the...
Hdud IM, Mobasheri A, Loughna PT.Chondrocytes are regularly exposed to load-induced stimuli and have the capability to sense and respond to applied mechanical stress. However, the mechanisms involved in chondrocyte mechanotransduction are not clearly understood. The purpose of this study was to explore the effects of cyclic equibiaxial mechanical stretch on the expression of α-BK and TRPV4 channels. Results: Freshly isolated equine articular chondrocytes were subjected to mechanical stress (8% elongation at frequency of 0.5 Hz for 8 h). Western blotting was used to investigate the expression of BKCa and TRPV4 channel prote...
Rathelot J, Canioni P, Bosc-Bierne I, Sarda L, Kamoun A, Kaptein R, Cozzone PJ.Porcine and equine colipases have been submitted to mild tryptic digestion. Proteolysis occurs at the Arg5-Gly6 bond with the loss of the N-terminal pentapeptide. Studies of native and trypsin-treated colipases by circular dichroism and laser chemically induced dynamic nuclear polarization indicate that proteolysis induces conformational changes in the region of the tyrosine cluster. Experiments in the presence of phospholipid provide further evidence showing that these residues are in or close to the region of the protein interacting with aggregated lipids. Kinetic studies of the reaction of ...
Clarke LL, Argenzio RA.In contrast to in vivo findings, the equine proximal colon fails to demonstrate significant net absorption of Na+ and Cl- under in vitro conditions. The present study was undertaken to determine if endogenous prostanoids are responsible for this apparent lack of ion transport. Proximal colonic tissues from ponies were preincubated in either normal Ringer solution or in Ringer containing 1 microM indomethacin and studied in Ussing chambers containing these solutions. Untreated colonic mucosa demonstrated negligible Na(+)-Cl- absorption in the basal state. In contrast, indomethacin-treated colon...
Watson ED, Stokes CR, David JS, Bourne FJ, Ricketts SW.Intrauterine infusion of 1 per cent oyster glycogen solution was used to induce acute endometritis in four genitally normal mares. Numbers of viable neutrophils recovered in uterine washings had increased by 1 h after infusion and remained elevated for at least 72 h. There was a significant correlation between numbers of viable neutrophils and total protein concentrations and between prostaglandin (PG)F and PGE2 concentrations in washings. There was also a significant relationship between concentrations of 15-keto-13, 14-dihydro PGF2 alpha in plasma and PGF in washings. Intrauterine concentrat...
Ahern BJ, Van Eps AW, Boston RC, Franklin SH.To determine the degree and ease of arytenoid abduction achieved with abaxial placement of a FASTak II suture anchor compared to 2 suture patterns with different directions of insertion-caudomedial to craniolateral and medial to lateral. Methods: Ex vivo experimental. Methods: Cadaveric larynges from 10 Standardbred racehorses. Methods: Each larynx was sequentially instrumented with all 3 arytenoid suture attachment in random order: (1) abaxial placement of a FASTak II suture, (2) caudomedial to craniolateral suture, and (3) medial to lateral suture placement. Each construct was abducted at 5N...
Le Jeune SS, Macdonald MH, Stover SM, Taylor KT, Gerdes M.Objective-To determine whether partial transection of the medial branch of the suspensory ligament (MBSL) alters equine third metacarpal bone (MC3) condylar surface strains and forelimb, distal joint angles in a manner consistent with promotion of lateral condylar fracture. Study Design-In vitro biomechanical experiment. Sample Population-Right forelimbs from 7 Thoroughbred horse cadavers. Methods-Lateral and medial MC3 condylar, dorsal and abaxial, bone surface strains and distal joint angles were measured both before and after partial transection of the MBSL during in vitro axial limb compre...
Singh S, Chopra M, Dilbaghi N, Manuja BK, Kumar S, Kumar R, Rathore NS, Yadav SC, Manuja A.Isometamidium hydrochloride (ISMM) is an effective drug for the treatment of trypanosomosis, but it causes local and systemic toxicity. Isometamidium hydrochloride has limited therapeutic index and exhibit considerable variation in their prophylactic activities. We developed a trypanocidal nanoformulation using ISMM and polymers sodium alginate/gum acacia to enhance the efficacy of the drug at lower doses, while minimizing undesirable side effects. It was characterized by transmission electron microscopy and infrared spectroscopy for evaluation of size, morphology, functional groups, etc. In v...
Clulow JR, Maxwell WM, Evans G, Morris LH.Duck and chicken egg yolk were compared for their protective effects against cold shock during the cryopreservation of stallion sperm in a lactose-EDTA-glycerol cryodiluent. Methods: A completely randomised design was used. Procedure Ejaculates from five stallions (n = 14 ejaculates) were split and diluted to either 20 or 200 x 10(6) sperm/mL in a lactose-EDTA extender containing either duck or chicken egg yolk. The extended semen was then frozen in liquid nitrogen. The percentage of sperm total motility and forward progressive motility were assessed before freezing and at 0 and 1 hr after tha...
Pérez MD, Puyol P, Ena JM, Calvo M.The interaction of sheep, horse, pig, human and guinea-pig whey proteins with fatty acids has been studied. Using gel filtration and autoradiography, it was found that sheep beta-lactoglobulin and serum albumin from all species had the ability to bind fatty acids in vitro. Sheep beta-lactoglobulin, isolated from milk, had approximately 0.5 mol fatty acids bound per mol monomer protein, and albumin from sheep, horse and pig contained approximately 4.5, 2.9 and 4.7 mol fatty acids/mol protein respectively. However, beta-lactoglobulin from horse and pig milk had neither fatty acids physiologicall...
Ponthier J, Teague SR, Franck TY, de la Rebière G, Serteyn DD, Brinsko SP, Love CC, Blanchard TL, Varner DD, Deleuze SC.Myeloperoxidase (MPO) is a pro-oxidant enzyme contained in and released by neutrophils during degranulation or after lysis. Post-thaw semen contains MPO and its concentration is associated with decreased sperm motility. Recently, MPO concentration in post-thaw semen was shown to be associated with the presence of non-sperm cells (NSC). The objective of this study was to evaluate the effect of a single-layer colloidal centrifugation before cryopreservation on NSC and MPO concentrations in equine semen. The experimental design consisted of freezing semen with or without previous centrifugation t...
Field HJ, Awan AR, de la Fuente R.The compound (S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine (HPMPA) had been previously shown to be highly effective in treatment of EHV-1 in a murine model for the equine disease. This paper describes the isolation of a series of mutants resistant to the drug. Resistance was demonstrated in cell culture and one mutant was tested in a murine model. The resistant mutant was pathogenic for mice; infectious virus was recovered from respiratory tissues and blood at levels similar to the parental virus. However, the mutant showed a significant degree of resistance in vivo, thus proving the viru...
Miragliotta V, Raphäel K, Lussier JG, Theoret CL.The development of exuberant granulation tissue, a situation that in some ways resembles the human keloid, compromises both the aesthetic and functional outcomes of wound repair in horses. To help elucidate the underlying molecular mechanisms the spatio-temporal expression of lumican (LUM) mRNA and protein for their potential contributions to tissue remodelling of body and limb wounds, was examined in an established experimental model. Expression was studied in intact skin and in samples of 1-, 2-, 3-, 4- and 6-week-old wounds of the body and forelimb. Temporal gene expression was determined b...
Bustos CP, Leiva CL, Gambarotta M, Guida N, Chacana PA.Equine salmonellosis is caused by several Salmonella serotypes, including Salmonella Newport, which cause enterocolitis and diarrhea. Treatment usually includes the administration of antibiotics. However, since multidrug-resistant Salmonella is commonly detected, alternative options to control the pathogen are needed. One of these options is the use of specific egg yolk antibodies (IgY) for passive immunotherapy. Thus, the aim of our work was to produce IgY antibodies against an equine S. Newport strain and assess their in vitro inhibitory activity. To this end, laying hens were immunized with...
Gobeil PA, Yuan Z, Gault EA, Morgan IM, Campo MS, Nasir L.Equine sarcoids are skin tumours of horses caused by infection with BPV-1 or 2. Maintenance and replication of the viral genome depend upon the viral proteins E1 and E2. We examined the effects of an E2 specific siRNA on E2 and E1 viral gene expression, viral load and cell growth in BPV-1 transformed sarcoid-derived cells. Transfection with E2-siRNA caused a reduction in E2 and E1 mRNA expression as well as viral load, growth inhibition and decreased anchorage-independent growth. siRNA treated cells showed significantly higher apoptosis rates than control cells. Thus sequence specific targetin...
Sayasith K, Doré M, Sirois J.The preovulatory rise in gonadotropins causes an expansion of the cumulus-oocyte complex, a process requiring the induction of several genes. The objectives of this study were to clone the equine tumor necrosis factor alpha-induced protein 6 (TNFAIP6), and investigate its regulation in equine follicles during human chorionic gonadotropin (hCG)-induced ovulation. The isolation of the equine TNFAIP6 cDNA revealed that it contains an open reading frame of 834 bp (including the stop codon), encoding a predicted 277 amino acid protein that is highly similar (91-93% identity) to known mammalian homo...
Koch DW, Schnabel LV.Tendon injuries are common in both veterinary and human clinical patients and result in morbidity, pain, and lost athletic performance. Consequently, utilizing naturally occurring injuries in veterinary patients as a comparative model could inform the development of novel therapies and increase translation for the treatment of human tendon injuries. Mesenchymal stem cells (MSCs) have shown considerable efficacy for the treatment of experimental and clinical superficial digital flexor tendon injury in the horse; however, the reinjury rate following treatment can remain high and MSC efficacy in ...
Koekemoer JJ, Dijk AA.Since protection against African horsesickness (AHS) is serotype-specific, rapid serotyping of AHSV is crucial to identify the correct vaccine serotype for efficient control of the spread of AHS outbreaks, especially when they occur in non-endemic regions. This paper describes the first one-day serotyping procedure that requires only a single RT-PCR and hybridization and which can identify multiple serotypes in mixed infections in one assay. The same region of genome segment 2 of all nine AHSV serotypes is amplified in a single RT-PCR. A universal primer set, designed to amplify the 5'-termina...
Garcia-Duitama J, Chayer B, Garcia D, Goussard Y, Cloutier G.Erythrocyte aggregation is a non-specific marker of acute and chronic inflammation. Although it is usual to evaluate this phenomenon from blood samples analyzed in laboratory instruments, in vivo real-time assessment of aggregation is possible with spectral ultrasound techniques. However, variable blood flow can affect the interpretation of acoustic measures. Therefore, flow standardization is required. Two techniques of flow standardization were evaluated with porcine and equine blood samples in Couette flow. These techniques consisted in either stopping the flow or reducing it. Then, the se...
Ioannou A, Lambrou A, Daskalakis V, Pinakoulaki E.The coordination of nitrite in myoglobin (Mb) has been characterized by resonance Raman spectroscopy and the frequencies of the nitrite bound to the heme Fe as well to the 2-vinyl have been computed by density functional theory (DFT) calculations. The DFT Natural Bond Orbital (NBO) analysis and the extensive isotope-labeling in the resonance Raman experiments indicate that NO (O1NO2) is bound to the heme Fe via O1. Based on the vibrational characterization of the reversible transition between low and high spin FeONO/2-nitrovinyl species, we suggest that the key step that triggers the spin-chan...
Hegre OD, Marshall S, Hickey GE.Plasma amine oxidase activities (benzylamine oxidase and spermine oxidase) were determined in the sera of a number of species of various ages. Benzylamine oxidase (BZO) activity, measured spectrophotometrically, was present in bovine, equine, and ovine species examined. Generally its activity in serum increased with the age of the animal. Spermine oxidase activity (SPO) was estimated by a bioassay of in vitro toxicity and did not necessarily correlate with BZO. Cytotoxicity in the presence of spermidine was found only in the sera of the ruminant species examined. Serum activity tended to rise ...
